Bioluminescence techniques are showing considerable promise in the development of sensitive, specific and automated biochemical analytical procedures. These developments should aid in the delivery of health care as has already been demonstrated by the use of the firefly bioluminescence assay techniques for urinanalysis and bacterial counting. Recent quantum yield studies of the bacterial bioluminescence reaction indicate its potential for analytical procedures, with one to two orders of magnitude increase in sensitivity over methods previously used. This proposal aims towards an understanding of the basic mechanism of this reaction and its application to specific assay techniques. Specifically it proposes to i) determine the interaction of all of the substrates with the luciferase, ii) to determine how the emitting chromophore is formed from the interaction of FMN and the luciferase, iii) to clarify the nature of the intermediates, and iv) to develop assay procedures of improved sensitivity for FMN, FAD, NADH, NADPH, aldehyde and NADH dehydrogenase and other enzymes that may be linked to the NADH dehydrogenase.